Murein hydrolase (N-acetyl-muramyl-l-alanine amidase) in human serum

An enzyme was identified in human serum which unlike lysozyme cleaved the amide bond between N-acetyl-muramic acid and l-alanine of the peptide side chain of the rigid layer (murein) of Escherichia coli. The N-acetylmuramyl-l-alanine amidase released all of the peptide side chains including those to which the lipoprotein is bound. A portion of the peptide side chains of the Micrococcus lysodeikticus murein was also hydrolysed from the polysaccharide chains. E. coli, M. lysodeikticus, Bacillus subtilis and Staphylococcus aureus were not killed by the amidase. Treatment of E. coli with EDTA or osmotic shock rendered the cells sensitive to the amidase and they were killed. Possible biological functions of the amidase are discussed.The enzyme was separated from lysozyme in human serum. Gel permeation chromatography indicated a molecular weight of the active enzyme of 82,000 while gel electrophoresis in the presence of sodium dodecyl sulfate revealed a molecular weight of 75,000. Thus, the enzyme probably consists of a single polypeptide chain. Incubation with neuraminidase rendered the amidase more basic suggesting the release of sialic acid residues. The modified glycoprotein disclosed an increased activity to murein. Enzyme activity was inhibited by p-chloromercuribenzene sulfonate and ethyleneglycol-bis(2-aminomethyl) tetraacetate (EGTA) at 1 and 0.2 mM concentration, respectively, whereas EDTA up to 5 mM was without effect. The amidase was also inactivated by agents that reduce disulfide bridges.     

Nitrogenase from Rhodospirillum rubrum Relation between ‘switch-off’ effect and the membrane component. Hydrogen production and acetylene reduction with different nitrogenase component ratios

Nitrogenase activity of ‘membrane-free’ extracts, produced from nitrogenstarved Rhodospirillum rubrum to which 4 mM NH⁺₄ had been added is only about 10% of the activity in the control. The activity could be restored to 80% by including the membrane component, earlier found to activate R. rubrum nitrogenase, in the reaction mixture. The relation between this ‘switch-off/switch-on’ effect and the function of the membrane component is discussed.Hydrogen production catalyzed by R. rubrum nitrogenase is also dependent on activation by the membrane component. Hydrogen production is inhibited by acetylene but the degree of inhibition is dependent on the nitrogenase component ratio. The strongest inhibition is achieved at low MoFe protein/Fe protein ratios. The $\text{ATP}\text{2 e}{}^{\mathrm{?}}$ values are 4–5 at the component ratios giving the highest activity and increase at high MoFe protein/Fe protein ratios. CO inhibits acetylene reduction but has no effect on the hydrogen production.     

Drug induced changes in cardio-vascular parameters in the Atlantic cod,Gadus morhua

Summary Ventral (VAP) and dorsal (DAP) aortic blood pressure, heart rate (HR) and cardiac output ( ) were recorded simultaneously in unanaesthetized Atlantic cod, and the effects of vasoactive drugs on the cardio-vascular parameters studied. Mean resting values for the parameters were VAP=4,39 kPa, DAP=2,49 kPa, HR=41 beats/min, and = 29,1 ml/min×kg. Adrenaline constricted the systemic vasculature, dilated the branchial vasculature and caused a decrease of HR and due to a cholinergic reflex. After atropine pre-treatment this reflex was abolished, and the effect of adrenaline on blood pressure enhanced. A small decrease in persisted after atropine, presumably reflecting the effect of an increased end-systolic afterload.Phenylephrine produced a weak increase in systemic vascular resistance, while isoprenaline lowered both systemic and branchial vascular resistance. The effect of isoprenaline is probably mediated by beta adrenoceptors in both vascular beds, since propranolol antagonizes the effect.Acetylcholine in low doses produces a drop in without affecting HR, while higher doses also stop the heart. There is no significant change in either branchial and systemic vascular resistance after acetylcholine.Abbreviations VAP mean ventral aortic blood pressure - DAP mean dorsal aortic blood pressure - TBPD trans-branchial blood pressure drop - HR heart rate - SV stroke volume - cardiac output (ventral aortic blood flow) - VR _g branchial vascular resistance - VR _s systemic vascular resistance     

Sympathetic nervous control of blood flow in the gills of the Atlantic cod,Gadus morhua

Summary The effects of sympathetic nerve stimulation, adrenaline and isoprenaline on the inflow pressure and efferent arterial and venous flow rates were studied in a cod gill preparation perfused at constant flow rate.The dominant effect of adrenaline was a reduced inflow pressure, accompanied by an increase in arterial flow and a decrease in venous flow. Isoprenaline also decreased the inflow pressure, but the changes in both outflow rates were small or absent.Sympathetic nerve stimulation gave arterial and venous flow changes comparable to the adrenaline effects, but the inflow pressure increased during nerve stimulation. Propranolol has little effect on the nerve responses, but phentolamine abolished or reversed the increase in inflow pressure, and also decreased or abolished the changes in outflow rates.The possible sites of action of the sympathetic fibres, and the distribution of adrenoceptors in the effector tissue is discussed. It is concluded that the main effect of sympathetic nerve stimulation is -adrenoceptor mediated, involving constriction of the arterio-venous pathway. The-adrenoceptor mediated control of total branchial vascular resistance may largely depend on circulating catecholamines.     

Cartilage proteoglycan aggregates. Electronmicroscopic studies of native and fragmented molecules

1. Proteoglycan aggregates from bovine nasal cartilage were studied by using electron microscopy of proteoglycan/cytochrome c monolayers. 2. The aggregates contained a variably long central filament of hyaluronic acid with an average length of 1037nm. The proteoglycan monomers attached to the hyaluronic acid appeared as side chain filaments varying in length (averaging 249nm). They were distributed along the central filament at an average distance of about 36nm. 3. Chondroitin sulphate side chains were removed from the proteoglycan monomers of the aggregates by partial chondroitinase digestion. The molecules obtained had the same general appearance as intact aggregates. 4. Proteoglycan aggregates were treated with trypsin and the largest fragment, which contains the hyaluronic acid, link protein and hyaluronic acid-binding region, was recovered and studied with electron microscopy. Filaments that lacked the side chain extensions and had the same length as the central filament in the intact aggregate were observed. 5. Hyaluronic acid isolated after papain digestion of cartilage extracts gave filaments with similar length and size distribution as observed for the central filament both in the intact aggregate and in the trypsin digests. 6. Umbilical-cord hyaluronic acid was also studied and gave electron micrographs similar to those described for hyaluronic acid from cartilage. However, the length of the filament was somewhat shorter. 7. The electron micrographs of both intact and selectively degraded proteoglycans corroborate the current model of cartilage proteoglycan structure.     

Bud formation inHydra: Inhibition by an endogenous morphogen

Summary From crude extracts ofHydra tissue a substance has been purified which prevents or retards the asexual reproduction by budding. The molecular weight is in the range of 300 to 1000 daltons. Inhibition of bud formation can be observed with concentrations equivalent to the extract from one hydra per 4 ml, that is, to a more than 10,000-fold dilution of the initial crude extract of a hydra. The purified inhibitor is active at a concentration of less than 10^–8 M.Most of the inhibitor present inHydra is bound to cells. Within the cells the substance is mainly bound to particulate structures which sediment at 10,000 g. Its concentration is highest in the hypostomal region and decreases in the direction of the tentacles and peduncle. A second, lower, peak has been found in the basal disc. Treatment of the animals with a toxic agent (nitrogen mustard) which depletes the animal of interstitial cells, nematocytes and nematoblasts excludes the possibility that the inhibitor is present to any great extent in these cells. In conjunction with cell separation experiments by centrifugation of fixed cells in suspension, these results indicate that nerve cells are the most likely sites of storage of the inhibiting substance, although epithelial cells are not excluded as sources for the inhibitor.     

Analysis of early stages of budding inHydra by means of an endogenous inhibitor

Summary Buds originate inHydra attenuata at a position 1/3 of the body length from the basal disc. The position with respect to the vertical axes is determined first and the position of the bud on the circumference of this budding region is specified later.Bud formation in hydra is reversibly prevented by pre-treatment with an inhibitor purified from hydra tissue (Berking, 1977). Some hours after the end of the treatment with the inhibitor, bud formation is resumed. From the starting or restarting point of development after the inhibitory treatment to the visible beginning of bud formation, 4 intermediary stages were distinguished on the basis of different responses to a second treatment with inhibitor. The pre0treatment is followed immediately by a period of maximal sensitivity to the inhibitor, which varies in length. At the conclusion of this phase the time interval required for the visible appearance of buds is fixed (12 h). In this and the following phase another application of inhibitor can cancel the entire preparatory process from the pre-treatment onwards. A transition to near complete resistance to inhibitor is the basis for defining a third phase. In a fourth phase, immediately before the evagination of the bud starts, the proesence of the inhibitor will again hinder the development. Upon removal of the inhibitor the suppressed buds will appear.     

The organization of macronuclear DNA sequences associated with C4A4 repeats in the polytene chromosomes of Stylonychia lemnae

Gene libraries of the micronucleus and the macronuclear anlagen of the polytene chromosome stage of Stylonychia lemnae were screened for internal C₄A₄ repeats. The number of these internal repeats was shown to be identical in both kinds of nuclei. Analysis of macronuclear sequences associated with C₄A₄ in the polytene chromosomes showed that several macronuclear DNA sequences are clustered. However, interspersed between short exons of one gene are located exons of several other genes, i.e. the exon of one gene is an intron for several other genes.by M. Trendelenburg     

Structure of adsorbed fibrinogen obtained by scanning force microscopy

It is shown that scanning force microscopy (SFM), operated in the attractive mode, can be used to obtain high resolution pictures of adsorbed fibrinogen molecules on solid surfaces, without the need for staining or special microscope grids. SFM also reveals the three-dimensional structure of the adsorbed molecules. Two forms of adsorbed fibrinogen are demonstrated on hydrophobic silicone dioxide surfaces; a trinodular about 60 nm long and a globular with about a 40 nm diameter. Polymeric networks formed after storage of the surface with adsorbed fibrinogen in PBS for 11 days are also shown. The SFM-results for the trinodular structure suggest the existence of loops or peptide chains extending outside the basic structure of the fibrinogen molecule.